The specificity of enzymes to a specific analyte renders them useful as probes for the detection of the presence of analytes in a liquid medium. A specific class of enzymes which has been proposed for the use in analytical biochemical methods are redox enzymes in which case the detection of the presence and concentration of an analyte in a medium may either be based on the measurement of the flow of charge resulting from the enzymatic redox reaction or on the accumulation of products obtained in the enzymatic redox reaction. In such methods an electrode is used which comprises an electrode material made for example of gold or platinum on which there are immobilized redox enzymes. A redox reaction involves the transfer of electrons from the enzyme to the analyte (in a reduction reaction) or vice versa (in an oxidation reaction) and if there is an electrical communication between the redox center of the enzyme molecules and the electrode material, there is an electrical charge flow which can serve as an indication of the presence of the analyte and the extent of charge flow may serve as an indication of the analyte's concentration. Alternatively, the determination may be based on the measurement of a product of the reaction.
The basic requirement in such electrobiochemical systems is to develop an electrical communication between the enzymes' redox center and the electrode material (Heller, 1990; Wilson et al., 1987). Such electrical communication may be established by immobilization of the redox enzymes in functionalized redox polymers (Degani et al., 1989; Gorton et al., 1990; Foulds et al., 1988) or by chemical modification of proteins with electron transfer mediators (Degani et al., 1988; Heller, 1992). Most of the presently developed electrobiochemical electrodes were utilized in an oxidative pathway. However, it was recently shown that immobilization of enzymes in bipyridinium functionalized polymers (Willner et al., 1990) or functionalization of proteins by bipyridinium components (Willner et al., 1991) gives rise to an electrical communication in photochemical systems which may be used in reductive routes.
It is the object of the present invention to provide an analytical electrobiochemical method for the determination of the presence of an analyte in both reductive and oxidative pathways.
It is furthermore the object of the present invention to provide an electrode for use in the above method.
It is still a further object of the present invention to provide a process for the preparation of such electrodes.